UMI-guided single locus sequence typing method for phylotyping Cutibacterium acnes from skin samples

Introduction Cutibacterium acnes is a dominant member of the human skin microbiota and displays substantial strain-level diversity with relevance for skin health and disease. However, accurate characterization of C. acnes lineages directly from skin samples remains challenging due to low biomass, host DNA contamination, and limitations of short-read sequencing. Methods Here, we present SLST-Seq, a culture-independent approach based on single-locus sequence typing (SLST), enabling strain-level profiling of C. acnes from low-input skin-strip samples. SLST-Seq adapts the LUMI-Seq® synthetic long-read sequencing technology to the C. acnes SLST marker, combining unique molecular identifier barcoding with de novo assembly to reconstruct full-length SLST sequences with high accuracy. Result Method performance was validated using single-isolate controls, defined genomic DNA mixtures, spike-in dilution series, and run-specific controls, demonstrating high specificity, quantitative accuracy across a wide range of target-to-background DNA ratios, and strong run-to-run reproducibility. Applied to skin-strip samples from healthy volunteers, SLST-Seq generated robust SLST profiles and revealed marked inter-individual variability, with donor-specific community structures largely conserved between face and back skin sites. Discussion Overall, SLST-Seq provides a sensitive and scalable framework for in situ analysis of C. acnes population structure and supports high-resolution studies of skin microbiome composition from challenging clinical samples.